Spot analysis for glycoprotein determination in the nanogram range.
A flexible method for glycoprotein determination with microliter-volumes using spot analysis ("Tüpfelprobe") on cellulose acetate layers is described. With glucose oxidase as an example of a glycoprotein and fluorescein isothiocyanate-labelled Concanavalin A, a sensitivity of 10 ng is reached; in combination with horseradish peroxidase 1 ng of glucose oxidase can be detected. A simultaneous determination of protein and glycoprotein with one single spot of 0.5, 1 or 2 microliters of a glycoprotein solution can be performed. The method is independent of many common external influences, e.g. dodecyl sulfate, Triton X-100, NP 40, mercaptoethanol and desoxycholate. Only pretreatment of the glycoprotein with urea decreases the sensitivity.
D002482:Cellulose / Q000031:analogs & derivatives; D003902:Detergents; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D005949:Glucose Oxidase / Q000032:analysis; D006023:Glycoproteins / Q000032:analysis*; D007202:Indicators and Reagents; D008722:Methods; D008832:Microchemistry; D011506:Proteins / Q000032:analysis*; D013861:Thiocyanates
D003902:Detergents; D005452:Fluoresceins; D006023:Glycoproteins; D007202:Indicators and Reagents; D011506:Proteins; D013861:Thiocyanates; C005062:acetylcellulose; D002482:Cellulose; D005949:Glucose Oxidase; D016650:Fluorescein-5-isothiocyanate